Studies in Early and Metastatic Breast Cancer The development of target-based anticancer drugs, such as protein kinase inhibitors, has become an attractive therapeutic strategy in oncology. We are currently studying the role of these compounds, specifically flavopiridol, a cyclin-dependent kinase (CDK) inhibitor, and Erlotinib, an epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TK), in the treatment of patients with metastatic breast cancer. Flavopiridol Flavopiridol is a synthetic flavone derived from rohitukine, a natural product isolated from a plant indigenous to India. Flavopiridol became the first CDK inhibitor to be tested in clinical trial and the first phase I study was performed at the NCI. There is preclinical evidence that flavopiridol has potent effects in vitro and antitumor activity in vivo in breast cancer. Several in vitro studies also demonstrate flavopiridol's ability to enhance apoptosis induced by chemotherapy, including taxanes, which are agents active against breast cancer. We conducted a phase I/II study of docetaxel followed by flavopiridol in patients with metastatic breast cancer. Our hypothesis is that the addition of flavopiridol to docetaxel will increase the objective response rate in the treatment of patients with breast cancer. Our primary objective is to determine the maximum tolerated dose (MTD) and dose-limiting toxicity (DLT) of docetaxel followed by flavopiridol. Secondary objectives include the determination of the activity as measured by objective response rate of the combination and the evaluation of cell cycle parameters in tumor and buccal mucosa biopsies before and after treatment with the combination. This study opened in September 2000 and closed in January 2003. Five patients were enrolled onto the original schedule with flavopiridol as a 72-hour continuous infusion. Docetaxel was administered as a 1-hour infusion at an initial dose of 60 mg/m2 followed in 24 hours by a 72-hour infusion of flavopiridol at 50 mg/m2/day every 3 weeks. Because three of these patients experienced myelosuppression (grade 4 neutropenia for 7 days), the study was amended to administer flavopiridol as a 1-hour infusion and to start at a lower dose of docetaxel. Six patients were enrolled onto the new schedule. The purpose of this study was to determine the maximum-tolerated dose and dose-limiting toxicities of docetaxel followed by flavopiridol in patients with previously treated metastatic breast cancer. The efffect of docetaxel and flavopiridol on Ki67, (phosphorylated retinoblastoma protein) Rb, and p53 was explored. A total of eleven patients with metastatic breast cancer were enrolled. Five patients received docetaxel followed by flavopiridol as a 72-hour continuous infusion. Three of these patients experienced dose-limiting toxicity of grade 4 neurotropenia. The other six patients received docetaxel followed by flavopiridol as a 1-hour infusion daily for 3 days. On this schedule, dose-limiting toxicity was grade 3 hypotension. There were two patients that sustained stable disease (72-hour flavopirodol) and one partial response (1-hour flavopiridol). The immunohistochemical evaluation for cyclin D1, p53, ki67/MIB1, bcl2, and apoptosis by TUNEL assay of paired tumor (when available) and buccal mucosa biopsy samples is being performed by the Laboratory of Pathology and with Dr. Xiaowei Yang in the Molecular Profiling Core. Preliminary immunohistochemical analysis of serial tumor and buccal biopsies revealed a decrease in MIB-1 in three out of the first five patients. There was a decrease in the MIB-1 proliferative index in biopsied samples (tumor and buccal mucosa) from a breast cancer patient with skin involvement post-treatment and a decrease in cyclin D1 expression in her tumor post-treatment. Erlotinib (OSI-774, TarcervaTM) Erlotinib is an orally active, potent, and selective inhibitor of the EGFR-TK. Significant growth delay was seen in a range of human tumor xenografts as well as tumor regressions in several tumor models exposed to Erlotinib. Breast cancer is associated with aberrant or overexpression of EGFR. Overexpression of EGFR was thought to be present in approximately 45% of breast cancers and be associated with disease progression and reduced survival, thus making it an ideal disease for treatment with EGFR-targeted therapy. We conducted a pilot study to evaluate EGFR signaling after treatment with oral Erlotinib in patients with metastatic breast cancer; the hypothesis was that a reduction in the phosphorylation of EGFR, ERK, and AKT (components of the EGFR pathway) would be detectable in tumor and normal surrogate tissue biopsies of breast cancer patients after exposure to Erlotinib. Because not much is known about the relationship between Erlotinib's biologic effect in tumor tissue and clinical activity, molecular and pharmacodynamic endpoints (specifically EGFR phosphorylation and downstream indicators of suppression of EGFR function) were determined. Our primary objectives were to determine whether a change in EGFR phosphorylation was detected in tumors and skin and buccal mucosa (as potential surrogate tissues) before and after treatment with Erlotinib, to determine whether a change in other parameters of signal transduction that are downstream of EGFR, namely ERK and AKT, occur in these tissues before and after treatment with Erlotinib, and to determine the assay reproducibility of assessing phosphorylation of EGFR, ERK, and AKT in biopsied samples before and after treatment with Erlotinib. Our secondary objectives were to obtain pharmacokinetic data and attempt to correlate it with a change in EGFR phosphorylation in tumor biopsies, to measure phosphorylated EGFR, ERK, and AKT in tumor tissues via protein microarray analysis, and to evaluate protein binding of Erlotinib via measurement of -1-acid glycoprotein levels.